Commercial Applications for Enzyme-Mediated Protein Conjugation: New Developments in Enzymatic Processes to Deliver Functionalized Proteins on the Commercial Scale

Abstract

The field of protein conjugation most commonly refers to the chemical, enzymatic, or chemoenzymatic formation of new covalent bonds between two polypeptides, or between a single polypeptide and a new molecule (polymer, small molecule, nucleic acid, carbohydrate, etc.). Due to the modest selectivity of chemical methods for protein conjugation, there are increased efforts to develop biocatalysts that confer regioselectivity for site-specific modification, thereby complementing the existing toolbox of chemical conjugation strategies. This review summarizes key advances in the use of enzymes to functionalize proteins with commercial relevance. The examples put forth have demonstrated value at the industrial level or show promising industrial potential in the laboratory.

Opportunities and challenges for combining chemo- and biocatalysis

“…These functionalized proteins can be designed to possess many attractive factors, such as increased shelf life, bioavailability or other pharmacokinetic properties. The drive for innovation of protein-based products has seen researchers focus on functionalizing proteins with both small molecules and macromolecules 99 . These few examples of novel chemistries highlight the importance, as well as demonstrating the results that can be achieved on successful combination of catalysis concepts from different disciplines in a synergistic fashion.…”

Synthesis of CeO2 nanoparticles with different morphologies and their properties as peroxidase mimic

“…Catalyst plays an important role in all kinds of catalytic reactions. As the catalyst, natural enzyme is more ideal for its significant advantages including high catalytic efficiency, substrate specificity, and mild catalytic conditions . However, enzymes have the problems of high production and application costs.…”

Post‐translational Assembly of Protein Parts into Complex Devices by Using SpyTag/SpyCatcher Protein Ligase

“…As this method relies on sortase catalysis, experimental conditions such as enzyme concentration can be limiting. Recent reviews describe methods to enzymatically conjugate proteins, such as by using ligases, oxidoreductases, transferases and peptidases …”

“…Covalent protein‐coupling methods such as using transglutaminases, which promiscuously crosslink proteins between lysine and glutamine side chains, have also been used for protein ligation . The use of more specific microbial transglutaminases has increased the utility of this method for protein conjugation, and it has been adopted for use in a variety of commercial applications . Large protein fusions such as HaloTag, SNAP‐Tag, and CLIP‐tag have previously been used for multivalent protein assembly, although the large size of tags can provide steric hindrance during protein ligation or epitope binding on the cell surface …”

“…Recent reviewsd escribe methods to enzymatically conjugatep roteins,s uch as by using ligases, oxidoreductases, transferases and peptidases. [14][15][16][17] Split-inteins,w hichc ontain an N-terminal fragment (Int N ) and aC -terminal fragment (Int C )c an be fused to target proteins. Once reconstituted in solution, split inteins will spontaneously splice themselves out, and in the process, covalently Exploiting the innate modularity of proteins hasa llowed advancesa cross the fields of syntheticb iology and biotechnology.B yu sing standardized protein components as building blocks, complex, multiprotein assemblies with sophisticated functions can be generated;f eats previously not possible with strictly genetic-engineering approaches.…”